Lysophosphatidic acid activates NF-kappaB in fibroblasts. A requirement for multiple inputs

Lysophosphatidic acid (LPA) is a growth factor that exerts a number of well characterized biological actions on fibroblasts and other cells. In the present study, we investigated the possibility that LPA activates the transcription factor NF-kappaB. NF-kappaB is a target of cytokines, but its activation by other classes of agonists has raised considerable interest in the control of processes such as inflammation and wound healing through varied mechanisms. We find that LPA causes a marked activation of NF-kappaB in Swiss 3T3 fibroblasts as determined by the degradation of IkappaB-alpha in the cytosol and the emergence of kappaB binding activity in nuclear extracts. The EC50 for activation of NF-kappaB is 1-5 microM, a range similar to that reported for reinitiation of DNA synthesis and activation of the serum response element. Activation of NF-kappaB is attenuated by pertussis toxin and inhibitors of protein kinase C, and it is completely blocked by the Ca2+ chelator BAPTA-AM. The combination of phorbol ester and thapsigargin promotes an activation comparable with that of LPA. Activation by LPA is additionally inhibited by tyrphostin A25 but not genistein or AG1478, indicating a selective utilization of protein-tyrosine kinases, and by certain antioxidants, implying a role for reactive oxygen species. The activation is also inhibited by tricyclodecan-9-yl-xanthogenate (D609), implying a requirement for hydrolysis of phosphatidylcholine. The data demonstrate the utilization of multiple pathways in the activation of NF-kappaB by LPA, not inconsistent with the relevance of several families of GTP-binding regulatory proteins.